Statins are currently the most widely used lipid-lowering drugs because they reduce the incidence of hard cardiovascular end-points (cardiovascular death, myocardial infarction and stroke) by 25% to 35% in different patient populations. These populations include those with stable or unstable coronary artery disease, diabetics, and hypertensive patients with other risk factors. In general, statins are well tolerated although muscular, liver and gastrointestinal side effects can occur. Statins can be associated with a wide range of muscular side effects, from non-specific or atypical myalgias, to myopathy and the full-blown rhabdomyolysis syndrome.
Myalgias are defined as muscle pain or complaints of sore muscles that can either be generalized or localized. Such symptoms occur in up to 10% of patients and can force physicians to reduce dose, switch to another statin using a trial-and-error approach or stop the medication completely. These muscular symptoms can also contribute to the relatively high rate of patients stopping statin therapy within the first two years of the treatment. Thus, even the more benign muscular symptoms can have important consequences and limit the large clinical and socio-economic benefits potentially offered by these agents.
Myopathy, while less devastating than rhabdomyolysis, can also occur after treatment with statins and is defined as muscle pain and/or weakness with increased creatine kinase (CK) levels at least 10 times the upper limit of normal. The incidence of myopathy is approximately 1-5%. The known predisposing risk factors for statin-related muscle toxicity include renal insufficiency, hypothyroidism, hereditary or acquired muscle diseases, history of muscle toxicity with another statin or a fibrate, concomitant use of a fibric acid derivative, alcohol abuse, clinical settings where increased plasma levels of statins could occur, as well as Asian ancestry.
Rhabdomyolysis is a rare event (well below 0.1% of statin users) but constitutes a life-threatening condition characterized by severe muscle toxicity, large increase in plasma creatine kinase (CK) levels (exceeding 10,000 U/L) and renal insufficiency secondary to myoglobin toxicity. Rhabdomyolysis has caused several patient deaths and has led to the withdrawal of one statin from the market, cerivastatin, (Baycol, Bayer). The incidence of rhabdomyolysis was also recently shown to be increased with another HMG-CoA reductase inhibitor, simvastatin (Zocor, Merck & Co.), when administered at a high dose (A to Z trial).
Currently plasma/serum Creatine kinase (CK) measurement is used as a biomarker for statin-induced muscle toxicity. For the vast majority of cases, CK measurement remains uninformative despite the presence of symptoms. Plasma/serum CK is an unspecific marker because it can be elevated for many other reasons, including physical exercise. An even greater limitation is its poor sensitivity, since it becomes indicative only after a substantial damage to muscle cells involving CK leakage to plasma from tissues. Thus, to this end it is well justified to develop new biomarkers for diagnosis of statin-induced muscle toxicity. Earlier studies (Phillips P S et al.: “Statin-associated myopathy with normal creatine kinase levels.” Ann Intern Med. 2002 Oct. 1; 137(7):581-5) on muscle specimens obtained from patients during acute muscle pain have demonstrated, e.g., an accumulation of inflammatory cells in histopathological studies.
The number of lipid mediators in the human body is overwhelming. Attempts have been made to facilitate their identification and quantification by advances in mass spectrometry and lipid biochemistry, which today enable the simultaneous high throughput identification and quantification of hundreds of molecular lipid species in several lipid classes (Ejsing C S, et al: Global analysis of the yeast lipidome by quantitative shotgun mass spectrometry. Proc Natl Acad Sci USA 2009, 106:2136-2141; Stahlman M, et al: High-throughput shotgun lipidomics by quadrupole time-of-flight mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 2009 Hiukka A, et al: ApoCIII-enriched LDL in type 2 diabetes displays altered lipid composition, increased susceptibility for sphingomyelinase, and increased binding to biglycan. Diabetes 2009, 58:2018-2026; Linden D, et al: Liver-directed overexpression of mitochondrial glycerol-3-phosphate acyltransferase results in hepatic steatosis, increased triacylglycerol secretion and reduced fatty acid oxidation. FASEB J 2006, 20:434-443.) collectively referred to as the lipidome. Lipidomic studies have sought to identify lipid cellular distribution and to describe their biochemical mechanisms, interactions and dynamics. Lipidomics is capable in principle of quantifying the exact chemical composition of lipidomes (Han X, Gross R W: Global analyses of cellular lipidomes directly from crude extracts of biological samples by ESI mass spectrometry: a bridge to lipidomics. J Lipid Res 2003, 44:1071-1079).
The bulk of the lipid data in the art today presents lipids in a sum composition format, i.e., phosphatidylcholine (PC) 34:1 (Brugger B, et al: Quantitative analysis of biological membrane lipids at the low picomole level by nano-electrospray ionization tandem mass spectrometry. Proc Natl Acad Sci USA 1997, 94:2339-2344) where the molecular lipid and the attached fatty acid tails remain unidentified. The identification of molecular lipid species, e.g., PC 16:0/18:1 (Ekroos K, et al: Charting molecular composition of phosphatidylcholines by fatty acid scanning and ion trap MS3 fragmentation. J Lipid Res 2003, 44:2181-2192) is the main feature of advanced lipidomics, which delivers highly resolved molecular lipid species rather than summed fatty acid information. For example, the information of the type of fatty acids and their positions of attachment to the glycerol backbone making up the particular PC molecule is revealed. There are conventional techniques such as thin-layer chromatography combined with gas chromatography but they not only require considerably larger sample amounts and laborious sample preparation, but they do not deliver the molecular lipid species. Despite multiple mass spectrometry techniques capable of characterizing lipid entities, most of them are still unable to deliver reliable high-quality quantitative data in terms of absolute or close-to absolute concentrations.
There is a need for specific and reliable methods for the detection and diagnosis of statin-induced muscle toxicity, as well as markers useful in this regard. There is also a need for improvements of existing treatment regimes with statins or lipid lowering drugs.